To determine whether this reciprocal regulation of Notch and Ikaros occurs in human T-ALL, we expressed active intracellular Notch receptors in human T-ALL cell lines and showed that Notch activation does not significantly increase the expression of non-DNAbinding Ikaros isoforms. While Notch activation has little effect on the pattern of Ikaros isoform expression in human T-ALL cell lines, Ikaros exerts a repressive effect on Notch signaling, similar to the Notch repression described in murine T-ALLs. Specifically, we demonstrated that Ikaros overexpression inhibits growth of human T-ALLs and that Ikaros downregulates expression of Notch target genes Hes1 and Hes5 at the transcriptional level. Interestingly, Ikaros inhibits growth more effectively in the cell lines with stronger Notch activation. In addition, Ikaros was shown to inhibit the Notch target gene Hes1 in both Notch dependent and independent manners. These data support the hypothesis that disruption of Ikaros contributes to proliferation of human T-ALL through de-repression of Notch signaling. In summary, we have demonstrated that Ikaros represses Notch signaling in human T-ALL.